PROJECT SUMMARY The inability to acquire protective immunity against Plasmodium is a major obstacle to malaria control. In the prior funding period, we showed that the Plasmodium orthologue of the host cytokine macrophage migration inhibitory factor, termed PMIF, impairs germinal center formation and the differentiation of T follicular helper cells (Tfh), leading to a reduction in the development of memory B and T cells. We recapitulated genetic PMIF deficiency in a P. berghei strain by vaccination with a pmif RNA replicon, which delayed blood-stage patency, augmented Tfh cell and germinal center responses, and enhanced the differentiation of memory CD4 and liver-resident, circumsporozoite-specific CD8 T cells. We observed improved control of infection and complete protection from re-infection, which was replicated by adoptive transfer of CD8 or CD4 T cells from pmif RNA replicon immunized hosts. Our data indicate that Plasmodium parasites utilize PMIF, which shows strict evolutionary conservation, to interfere with immunologic memory. We propose the following Specific Aims to achieve our goal of better understanding ineffective immunity to malaria and providing a foundation for better vaccines: Aim 1. Define the impact and potential synergy of PMIF neutralization in the host response to circumsporozoite (CSP) vaccination. The partial and waning immunity observed with the CSP-based RTS,S vaccine is due in part to an inadequate memory T cell response. We hypothesize that combining CSP with PMIF vaccination will increase T cell memory to CSP and provide optimal and high-level protection. We will study this approach in collaboration with the Walter Reed Army Institute of Research by immunizing mice against PMIF together with the Falciparum Malaria Vaccine-013 (FMP013) in a human CSP-transgenic P. berghei mouse model. Aim 2. Define the impact of PMIF on Plasmodium liver-stage development. PMIF also supports initial sporozoite infection and liver-stage parasite development by survival signaling through the hepatocellular MIF receptor, CD74. We hypothesize that PMIF re-programs hepatocyte metabolism to inhibit the host-protective apoptosis of infected hepatocytes. We will elucidate the pathways involved and better define the impact of liver-stage PMIF expression on the developing immune response. Aim 3. Evaluate the therapeutic potential of a novel small molecule PMIF inhibitor. We have developed a first-in-class small molecule, 26k, that selectively blocks PMIF signaling through the host MIF receptor CD74, reduces liver-stage parasite burden, and fully protects mice from cerebral malaria. We hypothesize that pharmacologic PMIF antagonism with 26k may be a tractable approach for prophylaxis and liver-stage treatment, and may augment immunologic memory responses. Closer knowledge of the PMIF pathway will lead to better strategies for malaria therapy and vaccine development, and potentially may be generalized to other parasites that express orthologous MIF proteins.